1. Field of the Invention
This invention relates generally to the field of immunology and specifically to human monoclonal antibodies which bind and neutralize respiratory syncytial virus (RSV).
2. Description of Related Art
RSV is the major viral-pathogen of the pediatric respiratory tract and has been identified as a leading cause of pneumonia and bronchiolitis. In the United States alone, there is a relatively large population of infants and children, about 100,000 to 200,000, at high risk of developing severe or fatal RSV illness. The high risk population includes infants and children with bronchopulmonary dysplasia, congenital heart disease, cystic fibrosis, cancer or various forms of immunodeficiency, as well as adults immunosuppressed prior to bone marrow transplantation, for example (McIntosh and Chanock (1990) Virology, 2nd edn. (Fields and Knipe, eds) Raven Press, Ltd., New York, pp. 1045-1072).
Several lines of evidence indicate that antibodies mediate resistance to RSV infection and illness. First, there is a correlation between levels of maternal IgG antibodies to RSV and the resistance of infants to infection during the first months of life when the risk of severe disease is greatest (Ogilvie, et al., J. Med. Virol., 7:263, 1981). Second, pooled human IgG containing a high level of RSV neutralizing antibodies or appropriate murine monoclonal antibodies that neutralize RSV efficiently can protect small animals from pulmonary infection when administered prophylactically and can reduce the titer of virus in the lungs of small animals and experimental primates at the height of RSV infection when administered therapeutically (Walsh, et al., Infection and Immunity, 43:756, 1984; Prince, et al., J. Virol., 55:517, 1985; Prince, et al., Virus Research, 3:193, 1985; Prince, et al., J. Virol., 63:1851, 1987; Hemming, et al., J. Inf. Dis., 152:1083, 1985). Third, a clinical study of pooled human IgG containing a high titer of RSV neutralizing antibodies has provided preliminary indications that these antibodies can exert a therapeutic effect on serious RSV disease in infants and young children (Hemming, et al., Antimicrob. Agnts. Chemotherap., 31:1882, 1987). Given this evidence, there is considerable interest in developing neutralizing antibodies to RSV for immunoprophylaxis and therapy for protecting infants at high risk of serious disease and for therapy in cases of serious RSV lower respiratory tract infection.
At present, there is no RSV vaccine available. The strategy currently being evaluated for prophylactic efficacy entails periodic intravenous inoculation of human IgG prepared from pooled plasma. Because of the large quantity of globulin required (1 to 2 gm per kg) and the need to administer this material intravenously in the clinic or hospital over a 2 to 4 hour interval every month during the fall, winter and early spring, this strategy is not very practical.
The main neutralization antigens on the surface of the RSV virion are the major glycoproteins F (viral fusion) and G (attachment). Monospecific and serum prepared against immunoaffinity purified F or G glycoprotein neutralizes RSV with high efficiency (Walsh, et al., J. Gen. Microbiol., 67:505. 1986). The antiserum to F, but not G. also inhibits fusion of RSV-infected cells to neighboring uninfected cells.
There is a need to develop human RSV antibody preparations with greater specific activity than the pooled human plasma preparations. A potentially effective solution to this problem would be the utilization of human monoclonal antibodies to RSV. RSV-specific monoclonal antibody, in contrast to polyclonal antisera, contains, by its very nature, a higher concentration of specific antibody. Therefore, the use of monoclonal antibody would decrease the amount of globulin required for prophylaxis or therapy by several orders of magnitude. As a consequence, an effective dose of monoclonal antibody could be administered intramuscularly (IM), rather than intravenously (IV) over a long period of time. Prophylaxis of infants at high risk could be accomplished IM at home, avoiding the need for hospital treatment for IV administration of antibodies. A reduction in the amount of globulin needed for therapy should also make it possible to treat patients with early mild RSV lower respiratory tract disease by administering antibodies IM in order to prevent hospitalization. In addition, aerosol therapy becomes feasible due to the increased specific activity of monoclonal antibodies, and accompanying decrease in therapeutic concentration necessary, coupled with increased therapeutic efficacy of such antibodies when introduced directly into the lungs. In fact, for aerosol application, F(abxe2x80x2)2 fragments of the RSV monoclonal antibodies are sufficient. Useful antibody preparations should also be capable of neutralizing a wide range of RSV isolates, including those of both antigenic subgroups A and B. The two subgroups, A and B, circulate simultaneously in the population in varying proportion at different times and are estimated to be 50% related in the F glycoprotein and 1-5% related in the G glycoprotein (McIntosh and Chanock, supra).
During the last several years, the efficiency of topical immunotherapy for RSV infection has been increased by two modifications of previous methodology First, a mixture of RSV F murine monoclonal antibodies directed at the major conserved neutralization epitopes on this glycoprotein was shown effective in topical immunotherapy of RSV infection in the cotton rat. Second, delivery of RSV polyclonal antibodies directly into the lungs in a small particle aerosol less than 2 xcexcm) was also effective therapeutically. The use of monoclonal antibodies should decrease the amount of IgG required for therapy by at least 2 orders of magnitude. In other studies in cotton rats, parainfluenza virus type 3 (PIV3) antibodies were also shown to be therapeutic when administered directly into the respiratory tract. The usefulness of topical immunotherapy is not limited to RSV. This approach likely will be effective for other respiratory viral pathogens whose pathogenic effects are also limited to the cells that line the lumen of the lower respiratory tract.
The present invention provides human monoclonal antibodies which bind and neutralize antigenic subgroups A and B of respiratory syncytial virus (RSV) and cell lines which produce these monoclonal antibodies. Also provided are amino acid sequences which confer neutralization function to the paratope of these monoclonal antibodies and which can be used immunogenically to identify other antibodies that specifically bind and neutralize RSV. The monoclonal antibodies of the invention find particular utility as reagents for the diagnosis and immunotherapy of RSV disease.
A major advantage of the monoclonal antibodies of the invention derives from the fact that they are encoded by a human polynucleotide sequence. Thus, in vivo use of the monoclonal antibodies of the invention for diagnosis and immunotherapy of RSV disease greatly reduces the problems of significant host immune response to the passively administered antibodies which is a problem commonly encountered when monoclonal antibodies of xenogeneic or chimeric derivation are utilized.
The antibodies of the invention are particularly efficacious in ameliorating RSV disease when administered directly to the lungs. This was surprisingly found to be true of Fab fragments. Topical delivery of RSV antibodies directly into the lungs has a major advantage over parenteral administration of antibodies for therapy of RSV disease. Antibodies delivered by the former route are approximately 80 to 160 times more effective in therapy, thereby decreasing the amount of antibodies required for therapy by a factor of 80 to 160. A further reduction in amount of antibodies required for therapy can be achieved by using human monocional antibodies or xe2x80x9chumanizedxe2x80x9d murine monoclonal antibodies such that the amount required for therapy is reduced by an additional factor of 25 to 50. This means that the total amount of antibodies required for therapy by parenteral treatment can be reduced by a factor of 2000 to 8000 when monoclonal antibodies are administered directly into the lungs for treatment of RSV infection. The ability to utilize Fab fragments in vivo for respiratory viral infections provides significant advantages over the use of whole antibody molecules such as: (1) greater tissue penetration; (2) avoidance of effector functions associated with Fc, such as inflammation; and (3) rapid clearance.
The in vivo therapeutic effectiveness of Fab fragments in treating respiratory viral infection is surprising and unexpected in view of the fact that: (1) Fab""s are non-covalent and can only attach to a single site, thereby precluding cross linking on separate virus particles, which is commonly thought to be necessary for viral neutralization; and (2) the Fc portion is thought to be needed in viral clearance in order to effect the complement cascade and antibody dependent cell cytotoxicity (ADCC). In view of the unexpected finding that pulmonary administration of Fab fragments which specifically bind RSV could be used effectively to ameliorate RSV infection, it is now possible to broadly apply this teaching to any viral infection where the in vivo viral growth occurs at the lumenal surface of the respiratory tract.